Four-leaf Stage Research Articles

The P2 protein of wheat yellow mosaic virus acts as a viral suppressor of RNA silencing in Nicotiana benthamiana to facilitate virus infection.

Wheat yellow mosaic virus (WYMV) causes severe viral wheat disease in Asia. The WYMV P1 protein encoded by RNA2 has viral suppressor of RNA silencing (VSR) activity to facilitate virus infection, however, VSR activity has not been identified for P2 protein encoded by RNA2. In this study, P2 protein exhibited strong VSR activity in Nicotiana benthamiana at the four-leaf stage, and point mutants P70A and G230A lost VSR activity. Protein P2 interacted with calmodulin (CaM) protein, a gene-silencing associated protein, while point mutants P70A and G230A did not interact with it. Competitive bimolecular fluorescence complementation and competitive co-immunoprecipitation experiments showed that P2 interfered with the interaction between CaM and calmodulin-binding transcription activator 3 (CAMTA3), but the point mutants P70A and G230A could not. Mechanical inoculation of wheat with in vitro transcripts of WYMV infectious cDNA clone further confirmed that VSR-deficient mutants P70A and G230A decreased WYMV infection in wheat plants compared with the wild type. In addition, RNA silencing, temperature, ubiquitination and autophagy had significant effects on accumulation of P2 protein in N. benthamiana leaves. In conclusion, WYMV P2 plays a VSR role in N. benthamiana and promotes virus infection by interfering with calmodulin-related antiviral RNAi defense.

Physiological and structural traits contribute to thermotolerance in wild Australian cotton species.

Five species of cotton (Gossypium) were exposed to 38°C days during early vegetative development. Commercial cotton (Gossypium hirsutum) was contrasted with four wild cotton species (G. australe, G. bickii, G. robinsonii and G. sturtianum) that are endemic to central and northern Australia. Plants were grown at daytime maxima of 30°C or 38°C for 25 d, commencing at the four-leaf stage. Leaf areas and shoot biomass were used to calculate relative rates of growth and specific leaf areas. Leaf gas exchange measurements revealed assimilation and transpiration rates, as well as electron transport rates (ETR) and carboxylation efficiency (CE) in steady-state conditions. Finally, leaf morphological traits (mean leaf area and leaf shape were quantified), along with leaf surface decorations, imaged using scanning electron microscopy. Shoot morphology was differentially affected by heat, with three of the four wild species growing faster at 38°C than at 30°C, whereas early growth in G. hirsutum was severely inhibited by heat. Areas of individual leaves and leaf numbers both contributed to these contrasting growth responses, with fewer, smaller leaves at 38°C in G. hirsutum. CO2 assimilation and transpiration rates of G. hirsutum were also dramatically reduced by heat. Cultivated cotton failed to achieve evaporative cooling, contrasting with the transpiration-driven cooling in the wild species. Heat substantially reduced ETR and CE in G. hirsutum, with much smaller effects in the wild species. We speculate that leaf shape, as assessed by invaginations of leaf margins, and leaf size contributed to heat dispersal differentially among the five species. Similarly, reflectance of light radiation was also highly distinctive for each species. These four wild Australian relatives of cotton have adapted to hot days that are inhibitory to commercial cotton, deploying a range of physiological and structural adaptations to achieve accelerated growth at 38°C.

A high-efficiency transient expression system mediated by Agrobacterium tumefaciens in Spinacia oleracea leaves

BackgroundOptimization of a highly efficient transient expression system is critical for the study of gene function, particularly in those plants in which stable transformation methods are not widely available. Agrobacterium tumefaciens‑mediated transient transformation is a simple and low-cost method that has been developed and applied to a wide variety of plant species. However, the transient expression in spinach (Spinacia oleracea L.) is still not reported.ResultsWe developed a transient expression system in spinach leaves of the Sp75 and Sp73 varieties. Several factors influencing the transformation efficiency were optimized such as Agrobacterium strain, spinach seedling stage, leaf position, and the expression time after injection. Agrobacterium strain GV3101 (pSoup-p19) was more efficient than AGL1 in expressing recombinant protein in spinach leaves. In general, Sp75 leaves were more suitable than Sp73 leaves, regardless of grow stage. At four-leaf stage, higher intensity and efficiency of transient expression were observed in group 1 (G1) of Sp75 at 53 h after injection (HAI) and in G1 of Sp73 at 64 HAI. At six-leaf stage of Sp75, group 3 (G3) at 72 HAI were the most effective condition for transient expression. Using the optimized expression system, we detected the subcellular localization of a transcriptional co-activator SoMBF1c and a NADPH oxidase SoRbohF. We also detected the interaction of the protein kinase SoCRK10 and the NADPH oxidase SoRbohB.ConclusionThis study established a method of highly efficient transient expression mediated by Agrobacterium in spinach leaves. The transient expression system will facilitate the analysis of gene function and lay a solid foundation for molecular design breeding of spinach.

Evaluation of Garlic Germplasm for Resistance to Leaf Blight Caused by Stemphylium vesicarium

Leaf blight caused by Stemphylium vesicarium is one of the most important fungal diseases of garlic (Allium sativum L.) worldwide, which results in a reduction of quality and yield. The breeding of resistant cultivars is an efficient approach to decrease the use of chemical fungicides and minimize crop losses. In this study, to find the resistant garlic resources against S. vesicarium, we evaluated the resistance degree of 20 garlic germplasms. To do this, garlic seedlings at four-leaf stage were rubbed with nonabsorbent cotton and then inoculated with spore suspension (3.0×105 spores/ml of potato dextrose broth) of S. vesicarium by spray method. Three to seven days after inoculation, the infected leaf area (%) of garlic seedling was measured. ‘Daeseo’ and ‘Namdo’ were included as susceptible and resistant control cultivars, respectively. After 3 to 7 days of incubation, the infected leaf area (%) of garlic seedling was measured. Our results showed that IT245512, IT245528, and IT244068 lines exhibited the highest resistance against S. vesicarium, whereas IT257134 and IT253043 lines were more susceptible than the susceptible cultivar ‘Daeseo’. Based on the results, the resistant genetic resources selected in this study can be used a basic material for resistant garlic breeding system against leaf blight.

Morphological and proteomic study of waterlogging tolerance in cotton

Floating seedling cultivation technique is a novel seedling method in cotton and it provides an ideal model to study cotton growing under waterlogging stress. Morphological character and proteomic profile of the primary root from the seedling cultured by the new technology were evaluated in this study. Compared to seedlings cultured by the traditional method, the diameter of the taproot from floating technology is small at all five seedling stages from one-leaf stage to five-leaf stage. There are similar changes between the thickness of cortex and diameter of stele, which increased from the one- to the two-leaf stage but decreased from the two- to the five-leaf stage. At the one-leaf stage, the number and volume of mitochondria in the primary root-tip cells were less than those in the control. At the two-leaf stage, there was significantly less electron-dense material in the primary root-tip cells than those in the control group. From the one- to the two-leaf stage, the vacuole volume was significantly smaller than that in the control. Total 28 differentially expressed proteins were revealed from aquatic and control group roots of cotton seedlings at the three-leaf stage by two-dimensional electrophoresis, which included 24 up-regulated and four down-regulated proteins. The relative expression of the phosphoglycerate kinase (PGK) gene in aquatic roots increased from the one- to the four-leaf stage but declined rapidly from the four- to the five-leaf stage. The relative expression of the 14–3-3b gene tended to decrease from the one- to the five-leaf stage. The PGK and 14–3-3b genes were specifically expressed in the aquatic roots at the three-leaf stage. In brief, these changes induced waterlogging resistance in the aquatic roots of cotton seedlings in the floating nursery, thereby causing the roots to adapt to the aquatic environment, promoting the growth and development of cotton seedlings.

Autotetraploidy of rice does not potentiate the tolerance to drought stress in the seedling stage

Polyploid is considered an advantage that has evolved to be more environmentally adaptable than its diploid. To understand if doubled chromosome of diploid rice can improve drought tolerance, we evaluated the diploid (2X) and autotetraploid (4X) plants of three indica and three japonica varieties. Drought stress in the plastic bucket of four-leaf stage revealed that the drought tolerance of 4X plants was lower than that of its diploid donor plants. The assay of photosynthetic rate of all varieties showed that all 4X varieties had lower rates than their diploid donors. The capacity for reactive oxygen species production and scavenging varied among different 2X and 4X varieties. Further, transcriptomic analysis of 2X and 4X plants of four varieties under normal and drought condition showed that the wide variation of gene expression was caused by difference of varieties, not by chromosome ploidy. However, weighted gene co-expression network analysis (WGCNA) revealed that the severe interference of photosynthesis-related genes in tetraploid plants under drought stress is the primary reason for the decrease of drought tolerance in autotetraploid lines. Consistently, new transcripts analysis in autotetraploid revealed that the gene transcription related with mitochondrion and plastid of cell component was influenced most significantly. The results indicated that chromosome doubling of diploid rice weakened their drought tolerance, primarily due to disorder of photosynthesis-related genes in tetraploid plants under drought stress. Maintain tetraploid drought tolerance through chromosome doubling breeding in rice needs to start with the selection of parental varieties and more efforts.

First report of pepper chlorosis-associated virus infecting tobacco (Nicotiana tabacum) plants in Sichuan Province in China.

Tobacco (Nicotiana tabacum) is an economically important crop in China, and more than 30 viruses have been reported to infect tobacco (Yin et al. 2022). In July 2022, we observed interveinal necrosis on tobacco leaves in fields in Sichuan Province (N 27.9172, E 105.6662) (Fig. 1). Total RNA was isolated from multiple leaves of one plant using an RNAprep Pure Polysaccharide Polyphenol Plant Total RNA Extraction Kit (TIANGEN, Beijing, China). Total RNAs were pooled, and a TruSeq Stranded Total RNA with RiboZero Gold Kit (Illumina, San Diego, CA, USA) was used to eliminate ribosomal RNA. An RNA-Seq library was constructed using VAHTS Universal V6 RNA-seq Library Prep (Nanjing Vazyme, China). High-throughput sequencing was performed on the Illumina DNBseq platform (BGI-ShenZhen, China), which yielded 20,102,087 reads with an average length of 150 nt (total size >6 Gb). Unaligned reads were assembled de novo using SPAdes (Bankevich et al. 2012). Contigs with length ≥200 nt were subjected to local BLASTn and BLASTx analyses against the GenBank nt and nr databases, respectively (Wang et al. 2022). A total of 23 contigs were identified through BLASTx (e-value cut-off = 10 -3), ranging from 631 to 1555 bp long, with 82% to 96% coverage to partial genomic sequences of pepper chlorosis-associated virus (PepCaV-Higashitsuno_2021; Accessions: LC719619 to LC719621) and one contig (6459 bp) with 99% similarity to tobacco mosaic virus (Accession: OP525281) isolate DSMZ PV-0109 from Germany. The complete genome sequence of PepCaV was obtained using primers based on the assembled contigs. The 5'- and 3'-terminal regions of the RNA genome were obtained by 5'- and 3'-rapid amplification of cDNA ends. These amplicons were cloned using the pEASY-Blunt Zero Cloning Kit (TRANSGEN, Nangjing, China) and sequenced by Sanger sequencing. Complete genome sequences of tripartite PepCaV from tobacco samples were 7697, 1808, and 1557 nucleotides long (Accession: OR451987 to OR451989) and showed genome organization typical of the genus Ophiovirus in the family Aspiviridae. The complete sequences of RNA1, RNA2 and RNA3 genome segments shared 92.36%, 90.43%, and 95.24%, nucleotide sequence identities, respectively, with the isolate PepCaV-Higashitsuno_2021 pepper isolate (Accession: LC719619 to LC719621) (Shimomoto et al. 2023), but PepCaV has not been reported to infect N. tabacum. In June 2023, 10 plants collected from each place of Macheng (N 27.9094, E 105.6740), Xiangyang (N 28.0936, E 105.6249) and Moni (N 27.8899, E 105.5936) showing interveinal necrosis symptoms were tested using RT-PCR using PepCaV-MP610-F (5'-TGTTCTCTGCTATGCGGTTG -3') and PepCaV-MP610-R (5'-AGCAATCTCGCACCTGAAGT-3') to product 610bp amplicon. Twenty-five tobacco plants were positive for PepCaV. Single sequence from each location was submitted to GenBank (Accession: PP728631 to PP728633). Sap extracts from the original field leaf samples collected from Sichuan Province were used to mechanically inoculate tobacco plants (10 plants) at the four-leaf stage. After 7 days, leaf samples were tested using RT-PCR assay specific to PepCaV and TMV while samples were positive only for TMV but failed to transmit PepCaV by mechanical inoculation. According to previous literature, ophioviruses may be transmitted though soilborne fungus (Jeong et al. 2014). Further research is needed to understand the transmission, epidemiology, and pathological properties of the PepCaV. To our knowledge, this is the first report documenting natural PepCaV infection of tobacco plants in China, providing a scientific basis for PepCaV infection control in tobacco plantations.

Assessing Interactions between Nitrogen Supply and Leaf Blast in Rice by Hyperspectral Imaging

Mineral nitrogen (N) supply reportedly increases rice susceptibility to the fungal pathogen Magnaporthe oryzae causing blast disease. These biotic and abiotic factors cause changes in spectral reflectance of leaves; however, the effects of N × pathogen interactions on spectral characteristics of rice have not been studied. In this study, hyperspectral imaging was used to assess the effect of N supply on symptoms of rice leaf blast under greenhouse conditions. Three rice genotypes differing in blast susceptibility grown at low, medium, and high N supply were inoculated at the four-leaf stage with three M. oryzae isolates differing in virulence. The reflectance spectra (400 to 1000 nm) of healthy and symptomatic leaves were analyzed using the spectral angle mapper algorithm for supervised classification. Mineral N supply increased the contents of chlorophyll and total N. The number and area of lesions and total blast severity varied depending on rice genotype—M. oryzae isolate interactions and the amount of mineral N applied. The reflectance spectra of healthy tissue and of blast symptom subareas differed with N supply; rice genotypes differed in the response to N supply. Infected plants at high mineral N supply could be distinguished from those at low N supply due to higher differences in the spectra of symptom subareas. Results reveal the potential (and limitations) of hyperspectral imaging for quantifying N effects on rice leaves, disease severity, and symptom expression. The impact of these findings on plant phenotyping and remote sensing under field conditions is discussed.

Influence of genotypes and crop growth stage at time of infection on seed transmission of Maize chlorotic mottle virus

Understanding the effect of seed transmission of Maize chlorotic mottle virus (MCMV) is important in managing the spread of the disease especially secondary transmission caused by vectors. This study was set out to investigate the effect of different genotypes and crop growth stage at the time of infection to rate of MCMV transmission from seeds to seedlings. The experiments were laid out in randomised complete block design with three replications. Detection of MCMV in the grow outs was determined using double antibody sandwich enzyme-linked immunosorbent assay and real time reverse transcription polymerase chain reaction. The results showed low rates of seed transmission from the genotypes in which their was detection of MCMV in their samples (0.08–1%). Similarly, crop growth stage at time of infection also showed low transmission rate of up to 0.1%, with highest transmission rate at the four-leaf stage.

A Combination of Three Genomic Regions Conditions High Level of Adult Plant Stripe Rust Resistance in Australian Wheat Cultivar Sentinel.

A seedling susceptible Australian common wheat cultivar Sentinel showed resistance to stripe rust under field conditions. A Sentinel/Nyabing3 (Nyb3)-derived recombinant inbred line (RIL) population was phenotyped. A DArTseq marker-based linkage map of the Sentinel/Nyb3 RIL population was used to determine the chromosomal location of the adult plant stripe rust resistance possessed by Sentinel. Three consistent quantitative trait loci (QTL); QYr.sun-1BL, QYr.sun-2AS and QYr.sun-3BS were detected, and they on an average explained 18%, 15.6% and 10.6% of the variation in stripe rust response, respectively. All three QTL were contributed by Sentinel. QYr.sun-1B corresponded to the previously characterized gene Yr29. Sentinel expressed resistance at the four-leaf stage at 21 ± 2 °C in the greenhouse. Monogenic segregation among the RIL population was observed when screened at the four-leaf stage at 21 ± 2 °C in the greenhouse, and the underlying resistance locus was temporarily named YrSen. QYr.sun-3BS peaked on YrSen. QYr.sun-2AS was mendelized by generating and phenotyping a mongenically sgregating F6 RIL population, and it was temporarily designated YrSen2. RILs carrying Yr29, YrSen and YrSen2 in combination exhibited responses like the parent Sentinel. Based on a comparison of the genomic locations and resistance expression with stripe rust resistance genes previously located in their respective chromosomes, QYr.sun-2AS (YrSen2) and QYr.sun-3BS (YrSen) were concluded to represent new loci.

Response of corn to reduced rates of tiafenacil applied at vegetative growth stages

Abstract Tiafenacil is a new nonselective protoporphyrinogen IX oxidase (PPO)–inhibiting herbicide with both grass and broadleaf activity labeled for preplant application to corn, cotton, soybean, and wheat. Early-season corn emergence and growth often coincides in the mid-South with preplant herbicide application in cotton and soybean, thereby increasing opportunity for off-target herbicide movement from adjacent fields. Field studies were conducted in 2022 to identify the impacts of reduced rates of tiafenacil (12.5% to 0.4% of the lowest labeled application rate of 24.64 g ai ha–1) applied to two- or four-leaf corn. Corn injury 1 wk after treatment (WAT) for the two- and four-leaf growth stages ranged from 31% to 6% and 37% to 9%, respectively, whereas at 2 WAT these respective ranges were 21.7% to 4% and 22.5% to 7.2%. By 4 WAT, visible injury following the two- and four-leaf exposure timing was no greater than 8% in all instances except the highest tiafenacil rate applied at the four-leaf growth stage (13%). Tiafenacil had no negative season-long impact, as the early-season injury observed was not manifested in a reduction in corn height 2 WAT or yield. Application of tiafenacil directly adjacent to corn in early vegetative stages of growth should be avoided. In cases where off-target movement does occur, however, affected corn should be expected to fully recover with no impact on growth and yield, assuming adequate growing conditions and agronomic/pest management practices are provided.

Informing the operation of intelligent automated intra-row weeding machines in direct-sown sugar beet (Beta vulgaris L.): Crop effects of hoeing and flaming across early growth stages, tool working distances, and intensities

The effective management of intra-row weeds is necessary to preserve crop yields in slow-to-establish direct-seeded crops like sugar beet, and suitable non-chemical control options are limited. However, the advent of automated intelligent intra-row weeding machines has provided a promising solution. These machines identify individual crop plants and selectively hoe or flame weed the between-crop zone, leaving the intra-row close-to-crop zone untreated. To characterize crop tolerance to intra-row weeding and describe optimal tool working intensities and distances, five pot experiments were performed using sugar beet as the test crop. From cotyledon through the four-leaf stage, hoeing could be implemented as close as 1 cm to the center of sugar beet plants before adverse crop effects were observed. Based on crop injury measured, direct flaming of sugar beet at the cotyledon stage is not advised at any intensity, although flaming tolerance was observed to increase with crop growth stage. Direct flaming did not affect sugar beets when the propane dose was ≤0.74 kg km−1 at the two-leaf stage, ≤1.49 kg km−1 at the four-leaf stage, and ≤5.95 kg km−1 at the six-leaf stage. Sugar beets at the two-leaf growth stage were not damaged by flaming at distances ≥1.5 cm at a propane dose of 3.72 kg km−1. Field experiments will be necessary to confirm these operational guidelines for next-generation automated intelligent weeders.

Synthesis, characterization, and evaluation of nanoparticles of clodinofop propargyl and fenoxaprop-P-ethyl on weed control, growth, and yield of wheat (Triticum aestivum L.)

Abstract Nanoherbicides are articulated by exploiting the prospective of nanotechnology for effectively delivering chemical and biological herbicides using nanomaterial‐based herbicide combinations. The nanoparticles were characterized using X-ray diffraction and FT-IR. On the targeted weeds, the nanoherbicides were sprayed at the third to fourth leaf stage. Six different doses were applied. The mortality and visual injury caused by both chitosan-based nanoherbicides reached 100% at the recommended dose of standard herbicide. The 5-fold lower dose exhibited weed density and maximum wheat yield and related parameters. For the same traits, the nanoherbicide at 10-fold lower dose of commercial herbicides showed a comparable influence as the suggested dose. The size of both herbicides was found to be 35–65 nm. It was observed that the clodinofop-propargyl nanoherbicide has an intense peak appearing at a 2θ value of 29.83°, corresponding to the (176) plane of the anatase phase and NPs of fenoxaprop-P-ethyl showed an intense peak around the 2θ value of 30.55° corresponding to the (74) plane of the anatase phase. The FT-IR spectra of fenoxaprop-P-ethyl clearly showed that the major functional groups were located in the FT-IR region between 610 and 1,840 cm−1 and the major functional ones of clodinofop propargyl were located in the FT-IR region between 640 and 1,740 cm−1. Nanoherbicides could restore the efficacy of conventional herbicides by improving stability and reducing toxicity.

Heterologous expression of heat-shock protein PpHSP70 improves high temperature and drought tolerance in rice

The research investigated the potential of heterologous expression of the heat-shock protein PpHSP70 in enhancing high temperature and drought tolerance in rice. After screening the whole genome sequence of Physcomitrium patens in the Phytzome database, 30 members of the HSP70 gene family were obtained. Their expression and the tissue expression patterns under abiotic stressors of high temperature, drought, and salt were analyzed. During the four-leaf stage of rice, overexpressed materials were subjected to a temperature of 45 °C for 20 h and to drought conditions for one week. After processing, 5 overexpression PpHSP70 materials survived against high temperature, and 7 materials survived against drought, while all wild-type materials KY131 died after the treatment. The results demonstrated that overexpression of PpHSP70 significantly improved both high-temperature tolerance and drought. This study provides valuable insights into the utilization of heterologous expression for enhancing crop tolerance to abiotic stressors.

First report of Stemphylium leaf blight of onion (Allium cepa) caused by Stemphylium vesicarium in Italy.

In July 2019, severe leaf symptoms were observed on onion plants (Allium cepa L. cv. Dorata di Parma) in a commercial field located in the municipality of Medicina (Bologna, Emilia-Romagna region), in northern Italy. Diseased leaves showed yellowish-pale-brown and oval-shaped lesions, which later coalesced in larger necrotic areas, and black leaf tips. As the disease progressed, conidia developed on the necrotizing leaves, until premature desiccation of the whole plants. Disease incidence of approximately 70% was calculated in the affected field, together with yield losses that were estimated to be above 30%. Symptomatic tissue fragments excised from the leaf lesions were surface disinfested with NaOCl 1% for 2 minutes, rinsed with sterile water and transferred onto potato dextrose agar (PDA). Fungi were consistently isolated after 5 days of incubation at 27 ± 1°C in the dark. Single spore isolation was performed on PDA to obtain 7 pure cultures, whose morphological characteristics were consistent with the description of Stemphylium vesicarium (Ellis 1971). DNA from a representative single spore isolate was extracted and the internal transcribed spacer region (ITS) of ribosomal DNA (rDNA) was amplified using the universal primers P-ITS1 and P-ITS4 (White et al. 1990). The PCR product was sequenced and deposited in GenBank (Accession No. OP144057). A BLAST search in CBS-KNAW collection bank (Westerdijk Fungal Biodiversity Institute, Utrecht, The Netherlands) showed 100% identity for the ITS gene with the strain of S. vesicarium under accession number CBS 124749. Moreover, the PCR assay using the primer pair KES 1999 and KES 2000 (Graf et al. 2016) for the cytochrome b gene displayed the specific fragments of 420 bp for S. vesicarium. The isolate was tested for pathogenicity on onion (potted plants cv. Texas Early Gran, fourth leaf stage) by spraying 4 ml of a conidial suspension (1 × 104 conidia/ ml) per plant. Inoculated and non-inoculated (sprayed with sterile distilled water) plants were kept at 24 ± 1°C and 90% relative humidity with a 16-h photoperiod. Seven days after inoculation, disease assessment was performed. Inoculated plants showed typical Stemphylium leaf blight (SLB) symptoms, similar to those observed in the field. No symptoms developed on the water-inoculated plants. S. vesicarium was consistently reisolated from the artificially inoculated onion plants and identified using a PCR assay, according to Graf et al. (2016). The assay was repeated twice with the same results. SLB is currently reported worldwide and it is considered a re-emerging threat and a truly challenging fungal disease, which can result in yield and quality losses of up to 90% in onion crops (Hay et al. 2021). In Italy, S. vesicarium has been reported several years ago on pear (Ponti et al. 1982) and, more recently, on radish sprouts (Belisario et al. 2008), chili pepper (Vitale et al. 2017) and spinach (Gilardi et al. 2022). To our knowledge, this is the first report of S.vesicarium on onion in Italy. Our results stress that development and implementation of innovative Integrated Pest Management (IPM) strategies are urgently needed to ensure an effective control of SLB, since only a few moderately resistant onion varieties are available (Hay et al. 2021) and no fungicides are currently registered to specifically control SLB in Italy. Further studies are underway to elucidate the pathogen geographic distribution and assess the impact of this disease on the onion crop in Italy.

First report of Fusarium tricinctum Species complex causing root rot of tobacco in China.

Tobacco (Nicotiana tabacum L.) is an economically important crop in China. In June 2021, a root rot disease was observed on tobacco (cv. Yunyan99) in Lushi, Mianchi, and Luoning counties of western Henan, China. Diseased tobacco plants exhibited wilting with leaf chlorosis and root rot accompanied by purplish to brown vascular discoloration. The symptoms were observed in four surveyed fields, 57 ha in total, and disease incidence ranged from 21 to 56%. Five symptomatic plants with leaf chlorosis and root rot were randomly collected from each field for pathogen isolation. Tissue pieces from diseased roots were surface sterilized in 75% ethanol for 30 s then rinsed with sterile distilled water three times, air dried, and placed onto potato dextrose agar (PDA) medium. Five isolates, SL1, SL2, SL3, LN and KC, were purified by single-spore culturing. On PDA, colonies grew at a rate of 2-5 mm/day and produced abundant cottony, white to pink aerial mycelia and rose pigment on the reverse side of the culture plate. From 7-day-old cultures grown on carnation leaf agar (CLA), macroconidia were straight to subarcuate, with blunt and slightly hooked apical and basal cells, had three to four septa, measured 23.4 to 44.6×3.5 to 4.2 μm (n=30). Cylindrical, napiform or oval microconidia were one to two-celled, measuring 6.3 to 22.9×2.2 to 4.9 μm (n=30). Spherical chlamydospores were intercalary or terminal, in chains. Such characteristics resembled those of the Fusarium tricinctum species comples (FTSC; Batra and Lichtwardt 1962; Leslie and Summerell 2006). To confirm the morphological diagnosis, the genomic DNA of the isolates were extracted, the translation elongation factor 1-alpha (EF-1α), RNA polymerase I largest subunit (RPB1) and second largest subunit (RPB2) genes were amplified with primers EF1/EF2, F5/G2R and 5f2/7cr respectively (O'Donnell et al. 2010), and sequenced. Maximum likelihood analysis was carried out using MEGA 7. Sequences were 97.55% to 100% identical to corresponding DNA sequences of FTSC based on GenBank and Fusarium MLST BLASTn analysis, and deposited in GenBank (ON637268.1-ON637272.1, ON637275.1-ON637279.1, ON637282.1-ON637286.1). Based on the morphological characteristics and phylogenetic analysis, the isolates were identified as F. acuminatum (SL1, SL2, SL3 and LN; Senatore et al. 2021) and F. reticulatum (KC; Moreira et al. 2019). Koch's postulates were conducted to verify the pathogenicity of individual isolates. The four-leaf stage healthy tobacco seedlings (Yunyan99, n=30) were inoculated by pouring 20 mL conidial suspension (1×107 conidia/mL) around the rhizosphere. Control seedlings were inoculated with sterilized water (n=30). All the treatments were maintained under greenhouse conditions with a 12-h light/dark photoperiod at 25±0.5℃ and 70% relative humidity for 30 days. The assay was conducted three times. Foliage chlorosis and root rot were observed on the inoculated tobacco seedlings, whereas the control seedlings remained asymptomatic after 30 days. The pathogens were reisolated from the necrotic tissue from all inoculated seedlings and were identified by sequencing partial EF-1α and RPB2 genes. Fusarium tricinctum species complex are known as an important causal of cereals Fusarium Head Blight (FHB; Laraba, et al. 2022). In China, F. acuminatum can also infect herb plants and fruits, such as Angelica sinensis, Schisandra chinensis (Ma et al. 2022; Shen et al. 2021). To our knowledge, this is the first report of root rot on tobacco caused by FTSC members in China as well as the world. This finding expands the host range known for FTSC and will be helpful for developing effective control strategies of tobacco root rot.

Water Deficit at Vegetative Stage Induces Tolerance to High Temperature during Anthesis in Rice.

Crop yields have been affected by many different biotic and abiotic factors. Generally, plants experience more than one stress during their life cycle, and plants can tolerate multiple stresses and develop cross-tolerance. The expected rise in atmospheric CO2 concentration ([CO2]) can contribute to cross-tolerance. Priming is a strategy to increase yield or to maintain yield under stress conditions. Thus, our objective was to evaluate if priming the rice plants with water deficit during the vegetative stage can induce tolerance to heat stress at anthesis and to evaluate the contribution of e[CO2]. The experiment was arranged in a completely randomized design in a factorial arrangement. Factor A consisted of the following treatments: water deficit at four-leaf stage (no-stress, and drought stress), heat at anthesis (normal temperature, high temperature), and priming with water deficit at four-leaf stage and heat stress at anthesis; and Factor B was two [CO2] treatments: a[CO2] = 400 ± 40 μmol mol-1 and e[CO2] = 700 ± 40 μmol mol-1. We assessed the effect of the treatments on plant growth, yield, biochemical, and transcriptome alterations. Although e[CO2] affected rice growth parameters, it did not affect the priming effect. Primed plants showed an increase in yield and number of panicles per plant. Primed plants showed upregulation of OsHSP16.9A, OsHSP70.1, and OsHSP70.6. These results showed induced cross-tolerance. Water deficit at the rice vegetative stage reduces the effect of heat stress at the reproductive stage. Water deficit at the vegetative stage can be used, after further testing in field conditions, to reduce the effect of heat stress during flowering in rice.

Unearth of small open reading frames (sORFs) in drought stress transcriptome of Oryza sativa subsp. indica

Drought is a major abiotic stress that influences rice production. Although the transcriptomic data of rice against drought is widely available, the regulation of small open reading frames (sORFs) in response to drought stress in rice is yet to be investigated. Different levels of drought stress have different regulatory mechanisms in plants. In this study, drought stress was imposed on four-leaf stage rice, divided into two treatments, 40% and 30% soil moisture content (SMC). The RNAs of the samples were extracted, followed by the RNA sequencing analysis on their sORF expression changes under 40%_SMC and 30%_SMC, and lastly, the expression was validated through NanoString. A total of 122 and 143 sORFs were differentially expressed (DE) in 40%_SMC and 30%_SMC, respectively. In 40%_SMC, 69 sORFs out of 696 (9%) DEGs were found to be upregulated. On the other hand, 69 sORFs out of 449 DEGs (11%) were significantly downregulated. The trend seemed to be higher in 30%_SMC, where 112 (12%) sORFs were found to be upregulated from 928 significantly upregulated DEGs. However, only 8% (31 sORFs out of 385 DEGs) sORFs were downregulated in 30%_SMC. Among the identified sORFs, 110 sORFs with high similarity to rice proteome in the PsORF database were detected in 40%_SMC, while 126 were detected in 30%_SMC. The Gene Ontology (GO) enrichment analysis of DE sORFs revealed their involvement in defense-related biological processes, such as defense response, response to biotic stimulus, and cellular homeostasis, whereas enriched Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways indicated that DE sORFs were associated with tryptophan and phenylalanine metabolisms. Several DE sORFs were identified, including the top five sORFs (OsisORF_3394, OsisORF_0050, OsisORF_3007, OsisORF_6407, and OsisORF_7805), which have yet to be characterised. Since these sORFs were responsive to drought stress, they might hold significant potential as targets for future climate-resilient rice development.

Soil and leaf phosphorus thresholds for modern potato production systems in tropical Oxisols

Precise diagnosis and prognosis of phosphorus (P) nutrition are essentials to improve P use efficiency and sustainability in potato cropping systems. Here, variable potato crop responses of 13 field experiments (2011–2022) were assessed to determine critical soil P-resin (SPresin) and potato leaf P (LP) concentrations to achieve maximum tuber yield in tropical Oxisols. The study comprised four experiments with the factorial design of potato cultivars in response to P fertilization rates and nine experiments with P fertilization rates as fixed effects. Relationships between initial SPresin test, LP measured at the third or fourth expanded leaf stage (i.e., 29–37 days after emergence, DAE), and relative total fresh tuber yield (RY) (yield of control/yield at highest P rate) were analyzed using linear-plateau, Mitscherlich, and quadratic-plateau regression models. A wide range of total fresh tuber yield (3.2–59 Mg ha-1) and LP (1.3–5.7 g P kg-1) were observed across experiments. The LP values in control plots was positively related with initial SPresin, reaching a maximum LP of 3.8 g P kg-1 at an initial SPresin of 80 mg P dm-3 (averaged across models). The relationship between RY and initial SPresin was significant in the three regression models (p < 0.05; R2 = 0.71–0.74). Maximum tuber yield was reached with the SPresin value of 93 mg P dm-3 (averaged across models). Across experiments and P fertilization rates, relationships between RY and LP were significantly described in the three regression models (p < 0.05; R2 = 0.55–0.67). The critical LP of 4.5 g P kg-1, obtained from the average across models, is proposed as the optimum indicator of potato P nutritional status to achieve maximum tuber yields in tropical Oxisols. These are the first critical P thresholds on a soil and environmental-specific conditions base available for tropical conditions in Brazil, timely informative in terms of optimization of P management to maximize tuber yield and P use efficiency, and minimize the risk of environmental P contamination.

Chitosan nanoparticles loaded with mesosulfuron methyl and mesosulfuron methyl + florasulam + MCPA isooctyl to manage weeds of wheat (Triticum aestivum L.)

Abstract Nanoherbicides are articulated by empowering the potential of nanotechnology for the efficacious delivery of chemical or biological herbicides with the aid of nanomaterial‐based herbicide combinations. Therefore, the goal of this work was to investigate the chitosan nanoparticles loaded with mesosulfuron methyl and mesosulfuron methyl + florasulam + (2-methyl-4-chlorophenoxyacetic acid) MCPA isooctyl herbicides as a possible environmentally benign substitute to manage weeds in wheat. Due to intriguing characteristics including biocompatibility, low allergenicity, biodegradability, and nontoxicity, chitosan biopolymers as sustainable chitin derivatives have received intense scrutiny in the biomedical business. The manufactured nanoparticles were characterized by using ultraviolet absorbance, scanning electron microscopy (SEM), X-ray diffraction (XRD), and Fourier transform infrared spectroscopy (FT-IR). The average particle size as revealed by SEM was 40–70 nm in a cluster form with the porous structure. The maximum absorption peaks of both nanoparticles of mesosulfuron methyl and mesosulfuron methyl + florasulam + MCPA isooctyl were 330 and 360 nm. The FT-IR analysis showed an intensive peak at 2θ value of 30.55° for mesosulfuron methyl and 32.79° for mesosulfuron methyl + florasula + MCPA isooctyl, which correspond to the 78 and 198 planes of the anatase phase, respectively. The nanoparticles were sprayed at the third to fourth leaf stages of the targeted weeds. Seven different doses were applied. A total of 100% mortality and visual injury were caused by the chitosan-based nanoparticles of both herbicides at the recommended dose of standard herbicide. The 5-fold lower dose showed the minimum chlorophyll content (5.75%), plant height (2.35 cm), fresh biomass (1.08 g), and dry biomass (0.33 g) of a weed mixture. For the same traits, the herbicide nanoparticles at 10-fold lower dose of commercial herbicides exhibited a similar effect as the recommended dose. Nanoherbicides could recuperate the conventional herbicide effectiveness by enhancing the stability and reducing the toxicity.